F12-23_DNA_ladder.jpgDNA Sequencing "Ladder" in an autoradioram

    Besides the ddNTPs, the DNA sequencing reaction formerly incorporated a radioactively - labelled dNTP. The radioactive label exposes a sheet of X-ray film laid on top of the transferred gel as an autoradiogram. The DNA sequence is read from bottom to top, according to the known order of the four termination reactions at top.

    The sequences of both DNA strands are often determined and compared as a check on accuracy. The sequence of the second strand will be the reverse complement of the first, which makes comparison tedious. Arrangement of the four reactions in the order shown allows the base sequence of the complementary strand to be read directly in the same order as the first strand, by turning the autorad over, and reading from top to bottom (think about it!).

    Homework: Read the sequence of this DNA ladder. Sequencing labs in Biol3250 was conducted like this, < 1991.

Figure © 2000 by Griffiths et al. ; text © 2007 by Steven M. Carr